title: Comparative analyses of the β-tubulin gene and molecular modeling reveal molecular insight into the colchicine resistance in kinetoplastids organisms.
abstract: Differential susceptibility to microtubule agents has been demonstrated between mammalian cells and kinetoplastid organisms such as Leishmania spp. and Trypanosoma spp. The aims of this study were to identify and characterize the architecture of the putative colchicine binding site of Leishmania spp. and investigate the molecular basis of colchicine resistance. We cloned and sequenced the β-tubulin gene of Leishmania (Viannia) guyanensis and established the theoretical 3D model of the protein, using the crystallographic structure of the bovine protein as template. We identified mutations on the Leishmania β-tubulin gene sequences on regions related to the putative colchicine-binding pocket, which generate amino acid substitutions and changes in the topology of this region, blocking the access of colchicine. The same mutations were found in the β-tubulin sequence of kinetoplastid organisms such as Trypanosoma cruzi, T. brucei, and T. evansi. Using molecular modelling approaches, we demonstrated that conformational changes include an elongation and torsion of an α-helix structure and displacement to the inside of the pocket of one β-sheet that hinders access of colchicine. We propose that kinetoplastid organisms show resistance to colchicine due to amino acids substitutions that generate structural changes in the putative colchicine-binding domain, which prevent colchicine access.
authors.affiliation: Laboratorio de Bioquímica y Biología Molecular de Parásitos, Instituto de Biología Experimental (IBE), Facultad de Ciencias, Universidad Central de Venezuela (UCV), Caracas 1041A, Venezuela.
authors.forename: Luis
authors.initials: L
authors.lastname: Luis
authors.forename: María Luisa
authors.initials: ML
authors.lastname: Serrano
authors.forename: Mariana
authors.initials: M
authors.lastname: Hidalgo
authors.forename: Alexis
authors.initials: A
authors.lastname: Mendoza-León
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003078:Colchicine / Q000494:pharmacology*; D004351:Drug Resistance / Q000187:drug effects*; D016830:Kinetoplastida / Q000187:drug effects* / Q000235:genetics*; D007891:Leishmania / Q000187:drug effects / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000187:drug effects; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D014404:Tubulin / Q000737:chemistry* / Q000235:genetics*
chemical_list: D014404:Tubulin; D003078:Colchicine